The PGC-1 coactivators increase the transcription of target genes;they have emerged as major regulators of many pathways involved in energy metabolism important in diabetes, hyperlipidemia and obesity. The liver functions as a central integrator of energy homeostasis, particularly the switch between the fed and fasted states. The previous cycle of this grant has funded studies showing that PGC-1 a plays a major role in expression of the genes of hepatic gluconeogenesis, while PGC-1[unreadable] is a positive activator of a broad program of hepatic lipogenesis and lipoprotein secretion. The present application proposes studies aimed toward moving these studies into a physiological realm, including studies of an interesting new genetic variant of PGC-1[unreadable] found in many humans. Specific Aim 1 suggests detailed physiological studies of mice with a liver-specific mutation in PGC-1a. We will study glucose and lipid homeostasis during fasting and feeding in these mice, we will also challenge these animals with high fat-induced obesity and insulin resistance. Detailed analyses of metabolism will include glucose and insulin tolerance tests as well as hyperinsulinemic-euglycemic clamps. Aim 2 will perform proteomic analyses of PGC-1a and PGC-1[unreadable] holo- complexes from murine liver, using a new isolation procedure we have developed. Importantly, we will study changes in the molecular anatomy of these complexes during physiological and pathophysiological states such as fasting, high fat feeding and obesity. Aim 3 will be focused on the mechanisms by which fatty acids stimulate the PGC-1[unreadable] promoter;preliminary data suggests an important role for the PPAR nuclear receptors. Finally, Aim 4 will focus on biochemical and genetic studies of a novel allele of human PGC-1[unreadable] (A203P) that has been associated with a reduced risk of obesity. This variant has reduced coactivation function on ERRa and LXRa, key coactivation partners of PGC-1[unreadable]. Analyses of this allele will include studies of its ability to activate gene expression in hepatocytes and liver in vivo. We will also knock this allele into the germ-line of mice, and study the effects of this polymorphic variant on hepatic energy metabolism, and energy homeostasis more systematically. These studies will allow us to analyze the role of this human polymorphism in obesity, and will also allow us to generate new hypotheses concerning the role of PGC-1[unreadable] in human disease.